The following description includes information that may be useful in understanding the present invention. It is not an admission that any of the information provided herein is prior art or relevant to the presently claimed invention, or that any publication specifically or implicitly referenced is prior art.
Autoimmune disorders, including neuroautoimmune diseases, affect 7-10% of the world population. Increasingly, such disorders are becoming associated with immune reactivity to commonly consumed foods. Ordinarily the gut mucosal immune system maintains immune homeostasis by inducing tolerance to antigens found in dietary proteins and peptides and commensal flora, while at the same time exerting immune defense against pathogens. The body's normal tolerance of “friendly” antigenic substances can, however, be disrupted by a number of factors. Intestinal barrier dysfunction and breakdown of gut-associated barriers can allow the entry of undigested proteins and peptides into circulation. Under these circumstances the ingestion of these food substances can result in the production of IgG and IgA antibodies, not only against the various food antigens but also against the body's own tissues (a phenomenon known as food autoimmune reactivity). This is due to homology (which is present to varying extents) between the amino acid sequences of many commonly consumed foods and those of many proteins that occur naturally in human tissue, including neural cells. As a result of this antigenic similarity or molecular mimicry between these various food proteins and different target tissue antigens, failing to detect food immune reactivities can initially result in the development of autoimmune reactivities and potentially lead to autoimmune (for example neuroautoimmune) diseases (Vojdani, 2014a; Vojdani, 2014b). As a result, food immune reactivities are receiving an increasing amount of attention, due to both their increasing prevalence and their adverse effect on health and quality of life (Johnson et al., 2014; Vojdani et al., 2014c).
All publications identified herein are incorporated by reference to the same extent as if each individual publication or patent application were specifically and individually indicated to be incorporated by reference. Where a definition or use of a term in an incorporated reference is inconsistent or contrary to the definition of that term provided herein, the definition of that term provided herein applies and the definition of that term in the reference does not apply.
The mechanism of immune reactivity is generally biphasic: an acute reaction occurs immediately following allergen exposure, followed by late phase reaction several hours later. During the acute reaction symptoms occur due to the binding of IgE and/or IgG to various cells and the release of mediators, such as histamine and platelet-activating factor (PAF), by mast cells, neutrophils and basophils The late phase involves the influx of inflammatory cytokines such as IL-4, IL-9, IL-33, and TNF-α, and cells such as neutrophils and eosinophils (Ho et al., 2012). In classical delayed food immune reactivity, production of high levels of IgG, IgM or IgA against various food antigens results in C1q binding to the antibody, with the formation of immune complexes and the deposition of immune complexes in various tissue sites. Symptoms can continue for days or even weeks following the initial immune reaction to such food antigens.
IgE, IgG, IgA or IgM play various roles in food immune reactivity (Mijayima et al., 1997). IgE functions via its high-affinity receptor, FcεRI, which is highly expressed on mast cells and basophils. IgG has several receptors, including the high-affinity FcγRI and FcγRIV receptors and the low-affinity FcγRIIB and FcγRIII receptors. All of these receptors are expressed on several types of cells involved in anaphylaxis, including mast cells, basophils, neutrophils, and macrophages.
Five different pathways are involved in food immune reactivity (Mancardi et al., 2013; Smit et al, 2011; Strait et al., 2002):    1. Classical pathway—involving IgE and its receptor FcεRI, mast cells and histamine    2. Alternative pathway—mediated by IgG1, FcγRIII, macrophages and the PAF pathway    3. IgG-basophil-PAF pathway    4. IgG-neutrophil-PAF pathway via FcγRIV    5. IgG, IgM or IgA-immune complex neutrophil pathway.All these reactions against dietary components can result from a failure of oral tolerance.
As noted above, the gut mucosal immune system normally maintains an immune homeostasis, which consists of maintaining tolerance to harmless or even beneficial molecules in the gut while mounting effective and appropriate immune responses against harmful pathogens (Lim and Rowley, 1982). A lack of response to food antigens with subsequent down-regulation of systemic immune response is what is characterized as oral tolerance. A failure in oral tolerance can result in immune reactivity to ingested food, with potentially life-threatening consequences such as allergies and autoimmunities (Tsuji and Kosaka, 2008).
When these different mechanisms of action fail to control ingested antigens, the result can initially be a breakdown in tolerance to soluble antigens, which activates secretory and systemic immune responses against food antigens. Individuals in whom the immune exclusion mechanism does not function can experience chronic hyperabsorption of macromolecules and the tendency to develop autoantibodies and even autoimmune disease (Maul and Dichmann, 2008). For this reason, the induction of IgG, IgM and IgA antibodies and immune complex formation to the actual food antigen and even cross-priming against bystander antigens may be of clinical significance. Both in vitro and in vivo experimental studies have demonstrated that IgG antibodies that are not balanced by a mucosal IgA response can enhance the epithelial penetration of bystander proteins (Brandtzaeg and Tolo, 1997). The passage of bacterial toxins and various food antigens through the epithelial cells can result in many immune disorders, including autoimmunities.
The type of systemic immune reaction against dietary proteins and peptides depends on the antigenic structure (e.g., protein antigens, particulate antigens, polysaccharides, glycoproteins, glycolipids or enzymes) and the genetic makeup of the individuals. For example, one person may produce IgG while another may produce IgA or IgM antibodies against dietary components (Barnes, 1995). If such IgG, IgM and IgA antibodies against dietary antigens are left undetected, the results can be the development of autoimmunity followed by autoimmune disease.
As a result, in recent decades significant progress has been made in the identification of target peptides in food antigens that share a similarity with autoantigens that are involved in autoimmune diseases (Baboonian et al., 1989; Baboonian et al., 1991; Lunardi et al., 1992; Lunardi et al., 2000; Ostenstad et al., 1995; Schrander et al., 1997). The glycine-rich cell wall protein peptide (GRP) represents an example of an antigenic peptide sequence that is able to prime T- and B-cell immune response in completely different and unrelated diseases. GRP is a ubiquitous food protein found in beans, fruits, vegetables and in gelatin. It has a very high degree of antigenic similarity/homology to ribonucleoprotein, fibrillar collagen, cytokeratin and EBV nuclear antigen-1 (EBNA-1) which are common antigens associated with autoimmune disorders.
This antigenic similarity between glycine-rich food antigen and Epstein-Barr virus and various tissue antigens involved in autoimmune disease can result in the production of cross-reactive antibodies. The finding of a common peptide epitope able to elicit an immune response in patients with food immune reactivities and different autoimmune disorders gives rise to the question of possible links between food antigens, gut mucosa, and systemic immune response (Lunardi et al., 1992; Schrander et al., 1997). Serum IgG antibodies directed against the GRP peptide were detected in several autoimmune disorders and in food allergic patients, and were able to cross-react with autoantigens including keratin, collagen and EBNA-1 (Lunardi et al., 2000). This data suggests that highly phylogenetically conserved epitopes in plants viruses and humans may be responsible for an autoimmune response in susceptible individuals. Furthermore, this indicates that the antigen spreading of a particular sequence between apparently divergent proteins may be involved in initiating or amplifying an immune response, resulting in autoimmunity in susceptible individuals.
An autoimmune response mediated by T-cell clones specific for particular food antigen epitopes can arise in the gut mucosa. Such T-cells can be recruited to particular sites, such as the joints, where they proliferate in response to homologous peptides derived from synovial proteins. Following local inflammation and up-regulation of MHC molecules, the release of additional self-antigens and/or epitope spreading can lead to a chronic, self-perpetuating process of organ inflammation and destruction resulting in autoimmunity (Lunardi et al., 1992; Vojdani, 2014a).
Recognition of food immune reactivity and associated health problems, particularly in regards to wheat and milk, has grown over the past two decades (Bousquet et al., 1998; Lack, 2008; Zuidmeer et al., 2008). A number of gluten peptides with the capacity to stimulate intestinal T-helper cells have been identified in celiac disease (CD) patients (Arentz-Hansen et al., 2000; Arentz-Hansen et al., 2002; Camarca et al., 2009; Tollefsen et al., 2006). A recent study showed that patients with non-celiac gluten sensitivity (NCGS) and Cohn's disease react to a repertoire of wheat antigens and produce IgG and IgA against them. This repertoire included various peptides, α-, γ-, ω-gliadins, glutenins, gluteomorphins and wheat germ agglutinin (Vojdani, 2011). Continuous exposure to environmental factors such as wheat not only causes NCGS and celiac disease but, if left untreated, can result in inflammation and autoimmunity (Counsell et al., 1994; De Freitas et al., 2002; Gillett et al., 2001). Indeed, celiac disease has been associated with various autoimmune disorders. The spectrum of autoimmune-associated antibodies detected in patients with CD or NCGS indicates that cross-reactivity and molecular mimicry occurs between gliadin and various tissue antigens (Alaedini et al., 2007; Collin et al., 2002; Frustaci et al., 2002; Hadjivassiliou et al., 2004; Jacob et al., 2005; Natter et al., 2001; Pratesi et al., 1998; Reinke et al., 2011; Vojdani et al., 2004).
Many studies have focused on the association between the prevalence of multiple sclerosis (MS) and dairy food consumption, and have found that the incidence of MS parallels the consumption of milk (Agranoff and Goldberg, 1974; Butcher, 1976; Kahana et al., 1994; Knox, 1977; Malosse et al, 1992). Notably, a high degree of sequence homology was found between a major protein of milk fat globule membrane called butyrophilin (BTN) and myelin oligodendrocyte glycoprotein (MOG) (Gardiner et al., 1992; Henry et al., 1999; Jack and Mather, 1990).
MOG (myelin oligodendrocyte glycoprotein) is a major antigen in the pathogenic autoimmune response of MS and its animal model, experimental autoimmune encephalomyelitis (EAE) (Vojdani et al., 2002). MOG is the only myelin autoantigen known to induce both a demyelinating autoantibody response and an ecephalitogenic CD4+ T cell response in animals with EAE (Amor et al., 1994). It has been found that an encephalitogenic T cell response to MOG can be either induced or, alternatively, suppressed as a result of immunological cross-reactivity (or “molecular mimicry”) with the extracellular IgV-like domain of the milk protein butyrophilin (BTN). In rats, active immunization with native BTN triggers an inflammatory response in the central nervous system characterized by the formation of scattered meningeal and perivascular infiltrates of T cells and macrophages (Vojdani et al., 2002). It has also been found that this pathology is mediated by an MHC class II-restricted T cell response of BTN that cross-reacts with MOG peptide sequence (Muthukumar M, et al., 2009).
Neuromyelitis optica (NMO) is a severe neuroautoimmune disorder that affects the gray and white matter in the brain and spinal cord, resulting in demyelination, axonal damage, and necrosis, and eventually resulting in paralysis and sensory loss in affected individuals (Jarius et al., 2008). In 75% of cases, NMO is associated with the presence of IgG1 antibody that binds selectively to aquaporin-4 (AQP4), which is a water channel belonging to the aquaporin family (Jarius et al., 2010; Kim et al., 2012). AQP4 is expressed in the astrocytic foot processes at the blood brain barrier, which are in contact with brain microvessels or subarachnoid space affecting solute concentration, electrical activity and modulation of neuronal transmission and excitability (Kinoshita et al., 2010). After binding, AQP4-specific IgG1 antibody has the capacity to first damage the astrocytes, and then cause demyelination in the spinal cord and optic nerve (Bradl and Lassmann, 2008). The binding of IgG1 to AQP4 also induces activation of the complement cascade and inflammatory infiltrates, which, after the induction of astrocytic cytotoxicity, cause demyelination and tissue destruction.
It has recently been suggested that pathogenic antibodies to AQP4 may be triggered by exposure to environmental proteins that have a similarity or molecular mimesis to a specific epitope of AQP4 (Vaishnav et al., 2013). Interestingly, spinach leaves express two thermally stable aquaporins that constitute 20% of the integral membrane protein (Plasencia et al., 2011). Similarly, soybean expresses aquaporins in germinating seeds as well as in the root nodules (Fleurat-Lassard et al., 2005). It has also been found that human AQP4 can cross-react with tomato and corn tonoplast intrinsic proteins (Vaishnav et al., 2013).
It has also been noted that an amino acid sequence with significant identity to a primary T-cell epitope in NMO occurs in a potentially immunogenic coat protein of the Parsnip Yellow Fleck Virus, which infects parsnips, celery, carrots, parsley, cilantro, chervil and dill. This epitope also shares significant sequence identity with a sequence present in a serine-protease inhibitor in the legume M. truncatula (Vaishnav et al., 2013).
It is apparent that many components of foods that have not yet been characterized can also have the potential to trigger autoimmunity. To date most studies associated with food immune reactivity have characterized only the water-soluble population of proteins and peptides present in the studied foods. An exception to this is wheat, as gluten (an alcohol-soluble component of wheat) has been used in food immune reactivity, cross-reactivity, and autoimmunity studies. In addition, the role of complement is not yet clear.
As noted above, to date food allergen studies have focused primarily on the detecting the presence of immunoglobulins to specific antigens, and have not addressed the issue of complement activation. United States Patent Application No. 2009/0010937 (to Chauhan) discusses detection of circulating immune complexes that include complement components such as C1q, however the methodologies that are discussed utilize cellular receptors for immune complexes to provide the immobilization necessary for detection using anti-complement antibodies. As such, they provide little to no insight into the antigen specificity of such complexes. U.S. Pat. No. 8,309,318 (to Dorval and Dantini) discusses the detection of allergen-specific immune complexes containing bound C3b using immobilized antigens. It has been demonstrated, however, that so called “innocent bystander” IgG-C3b adducts can form during complement activation (, which severely limits the utility of such an approach in determining the presence of antigen-specific complexes (Fries et al, 1984).
Thus there is a need for systems, devices, and methods for characterizing antibody (e.g. IgG, IgA, IgM, and other antibody classes) binding to a broader range of antigenic molecules found in food than are represented by water-extractable proteins and peptides in their natural state. In addition, there is a need for systems, devices, and methods that characterize the presence of complement components (e.g. C1q) associated with such antibodies.